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Background The complement cascade is a highly sophisticated network of proteins that are well regulated and directed in response to invading pathogens or tissue injury and forms a major component of our innate immune response. Complement C5a binds to two G-protein coupled receptors (GPCR); namely the C5a receptor (C5a1) and C5a receptor like-2 receptor (C5a2). C5a2 is a non-signalling GPCR and its role has been difficult to validate due to the unavailability of selective ligands. Aims Discover novel C5a2 ligands that allow us to probe and explore the functional role of C5a2 in cytokine release from macrophages. Methods Radioligand binding assays were used to screen a small peptide based library of 61 ligands designed to act at C5a1 for the ability to displace 125 I-C5a from C5a2 expressing membranes. A novel β -arrestin 2 recruitment via C5a2 BRET assay was developed to test for ligand activity at C5a2 and ligands were counter screened for β -arrestin 2 recruitment via C5a1. ELISA was used to measure cytokine release from human monocyte derived macrophages (HMDM). Results Radioligand binding testing of the 61 ligands revealed two ligands (P32 & P59) which showed ⩾ 40% inhibition of 125 I-C5a binding to C5a2 membranes at 100 μ M. P32 and P59 were able to dose dependently recruit β -arrestin 2 via C5a2 but not C5a1 each with an EC 50 of 7.6 μ M. Neither P32 nor P59 was able to recruit β -arrestin 2 to a similar level as C5a, with P32 able to recruit to ∼ 55% and P59 ∼ 30% of the level of C5a. Both P32 and P59 could dose dependently inhibit the release of IL-6 from HMDM when co-stimulated with LPS but had no effect on IL-10 release. Conclusions P32 and P59 are the first reported selective ligands for C5a2 and show novel pharmacology with the ability to modulate cytokine release from HMDM. [ABSTRACT FROM AUTHOR]

NLRP3 is a critical component of the inflammatory process and its aberrant activation is pathogenic in inherited disorders such as the cryopyrin associated periodic syndromes and complex diseases such as multiple sclerosis, type II diabetes and atherosclerosis. There is a compelling rationale to develop a potent, selective inhibitor of NLRP3. We demonstrate that a small molecule, MCC950, specifically blocks NLRP3 activation at nanomolar potency in response to numerous stimuli. MCC950 potently inhibits ASC oligomerisation, caspase-1 activation, IL-1 β secretion and cell death in response to NLRP3 but not AIM2, NLRC4 or NLRP1 inflammasome activation. MCC950 can block IL-1b in vivo and attenuates the course of the NLRP3 dependent, IL-1b driven disease model of multiple sclerosis, experimental autoimmune encephalomyelitis. Furthermore, MCC950 treatment rescues neonatal lethality in a murine model of cryopyrin associated periodic syndromes and is active in ex vivo samples from patients with Muckle–Wells syndrome. MCC950 is thus a novel potential therapeutic for NLRP3 associated syndromes and a useful tool for the further study of NLRP3 in human health and disease. [ABSTRACT FROM AUTHOR]